Evaluating Alfalfa Stress Tolerance

Project Title

Assessment of the Transcriptomic Response of Edited SPL8 Alfalfa Genotypes to Drought Stress.


Stacy Singer (AAFC Lethbridge) Stacy.singer@agr.gc.ca

Abdelali Hannoufa (AAFC London)

Status Project Code
In progress. Results expected in December, 2022 FRG.10.21


This team has successfully developed and used CRISPR/Cas gene editing technology targeting the SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE 8 (SPL8) gene in alfalfa. These edited plants are more drought tolerant in the greenhouse. But many genes have more than one job, so shutting off SPL8 may affect more than just drought tolerance. In this case, they suspect the edited plants may have different levels of gibberellic acid, which is involved in leaf size and plant height; an effect that is likely distinct from drought tolerance. They want to see what else might be affected in the edited plants that could be responsible for the observed drought resilience.


  • Compare global transcript levels (gene expression) in control and edited alfalfa genotypes under both well-watered and drought conditions to see what genes are affected when SPL8 is targeted using CRISPR/Cas.

What they will do

They will repeat the greenhouse drought experiment, harvest the appropriate tissues, extract total RNA and assess its quality, and send the samples for RNA sequencing (4 biological replicates of each). The large amount of data obtained will then be analyzed using bioinformatics in order to identify pathways and genes of interest that differ between edited and control genotypes.


The identification of particular genes that function downstream of SPL8 and are of importance for eliciting drought tolerance will further our ability to improve this trait in alfalfa and other forage species without any negative impacts on growth.